Benzimidazole vascular damaging agents

ABSTRACT

Vascular damaging agents composed of substituted 5(6)-substituted benzimidazole-2-carbamates are provided. These agents are useful in the preparation of medicaments for the treatment of diseases involving neovascularisation, particularly for the treatment of solid tumors, macular degeneration, diabetic retinopathy, rheumatoid arthritis, psoriasis, and atherosclerosis. Embodiments include a 5(6)-substituted benzimidazole-2-carbamate of formula I                    
     wherein A represents a multi-substituted alkyl group or aromatic ring.

This application is a 371 of PCT/GB00/00099 filed Jan. 14, 2000.

This invention relates to vascular damaging agents and particularly tothe use of new and known substituted benzimidazoles in the preparationof medicaments for the treatment of diseases involvingneovascularisation.

Formation of new vasculature by angiogenesis is a key pathologicalfeature of several diseases (J Folkman, New England Journal of Medicine333, 1757-1763 (1995)). For example, for a solid tumour to grow it mustdevelop its own blood supply upon which it depends critically for theprovision of oxygen and nutrients; if this blood supply is mechanicallyshut off the tumour undergoes necrotic death. Neovascularisation is alsoa clinical feature of skin lesions in psoriasis, of the invasive pannusin the joints of rheumatoid arthritis patients and of atheroscleroticplaques. Retinal neovascularisation is pathological in maculardegeneration and in diabetic retinopathy. In all these diseases reversalof neovascularisation by damaging the newly-formed vascular endotheliumis expected to have a beneficial therapeutic effect.

5(6)-Substituted benzimidazole-carbamates are known and have found useas antiparasitic agents (P. J. Islip in Burgers Medicinal Chemistry (M.E. Wolff ed.), Fourth Edition, Part II, p481, (1979)). Examples of suchcompounds include mebendazole, fenbendazole, oxibendazole, flubendazole,albendazole, cyclobendazole, parbendazole, dribendazole, luxabendazole,and etibendazole. Their mode of action for their antiparasitic action isbelieved to involve selective binding to tubulin of the target parasitewhile having little effect due to binding tubulin of the mammalian host(Biochim. Biophys. Acta 630, 271-278, (1980)). Some of these compoundshave been shown to be antimitotic for cancer cells and one particular5(6)-substituted benzirnidazole-2-carbamate, nocodazole, has thereforebeen studied as an anticancer agent (Cancer Research, 36, 905-916(1976)). No effects on neovasculature have been reported for any ofthese compounds.

Some structurally-unrelated compounds which bind tubulin have been shownto have anti-vascular effects when given at their maximum tolerated dose(MTD) (S. A. Hill et al. Eur. J Cancer, 29A, 1320-1324 (1993)) but othertubulin-binding agents, such as docetaxel, have no vascular-damagingactivity even when administered at the MTD. The presence oftubulin-binding properties is then not predictive for antivascularactivity.

According to the present invention there is provided the use of5(6)-substituted benzimidazole-2-carbamates for the preparation ofcompositions for the treatment of diseases involving angiogenesis inwhich the 5(6)-substituted benzirnidazole carbamate has the formula

wherein

Alk is an alkyl group

X is oxygen, sulphur, sulphinyl, sulphonyl, carbonyl (CO), thiocarbonyl(CS), sulphonyloxy, NH, iminomethylene (C═NH), N-hydroxyiminomethylene,N-alkoxyimninomethylene, dialkoxymethylene, 1,3-dioxolan-2yl,1,1-ethenyl, a group CHR³ or a bond

R¹ is hydrogen, alkylarninocarbonyl or alkoxycarbonyl

R² is hydrogen, alkoxycarbonyl, cyanomethyl, cyanoethyl, alkoxymethyl oracetoxymethyl.

R³ is hydrogen, hydroxy, alkoxy or amino

A is an optionally substituted aromatic, optionally substitutedheteroaromatic, optionally substituted heterocycloalkyl, optionallysubstituted alkyl or optionally substituted cycloalkyl group

and the pharmaceutically acceptable salts, solvates and hydratesthereof.

Particular substituents that may be present on the group A include oneor more substituents selected from a group Y, optionally substitutedalkyl,(where substituents on such alkyl group may include one or moreselected from hydroxy, amino, alkylamino, dialkylamino, halogen,carboxyl, SO₃H, sulphate, phosphate, alkoxycarbonyl, aralkoxycarbonyl,alkoxycarbonylamino, aminoalkylaminocarbonyl and cyano), halogen,hydroxy, amino, alkoxy, alkylthio, cyano, nitro, sulphate,isothiocyanate, aryl, heteroaryl and heterocycloalkyl.

Y is a group selected from phosphate, alkylphosphate, C(O)R⁴, OC(O)R⁴,SO₂R⁴, NHC(O)R⁴, NR⁵C(O)R⁴, SR⁴, S(O)R⁴, OSO₂R⁴, NHSO₂R⁴, NR⁵SO₂R⁴,SO₃H, CO₂H and CO₂R⁵ where R⁴ is a group selected from hydrogen, R⁵,OR⁵, NHR⁵, NR⁵R⁶, aryl, heteroaryl or heterocycloalkyl such aryl,heteroaryl or heterocycloalkyl groups being optionally substituted withone or more substituents selected from alkyl, heterocycloalkyl,haloalkyl, hydroxy, nitro, cyano, amino, alkylamino, dialkylamino,halogen, carboxyl, SO₃H, sulphate and phosphate. R⁵ and R⁶, which may bethe same or different, are each an alkyl group optionally substitutedwith one or more substituents selected from hydroxy, amino, alkylamino,dialkylamino, guanidino, halogen, carboxyl, SO₃H, sulphate, phosphate,aryl and heteroaryl.

Some of the compounds usable in the invention are known, for example thefollowing compounds within the following formula:

These compounds are

X A Fenbendazole S Ph Mebendazole CO Ph Albendazole S nPr Oxibendazole OnPr Nocodazole CO 2-thienyl

Certain of these compounds are novel. In one embodiment the novelcompounds are those of formula I in which at least one of thesubstituents on the group A is a group Y where Y is as hereinbeforedefined. Particularly preferred are compounds defined by the formula

wherein

alk is an alkyl group

B is an aromatic or heteroaromatic ring

X is oxygen, sulphur, sulphinyl, sulphonyl, carbonyl (CO), thiocarbonyl(CS), sulphonyloxy, NH, iminomethylene (C═NH), N-hydroxyiminomethylene,N-alkoxyiminomethylene, dialkoxymethylene, 1,3-dioxolan-2yl,1,1-ethenyl, a group CHR³ or a bond

R¹ is hydrogen, alkylaminocarbonyl or alkoxycarbonyl

R² is hydrogen, alkoxycarbonyl, cyanomethyl, cyanoethyl, alkoxymethyl oracetoxymethyl

R³ is hydrogen, hydroxy, alkoxy or amino

Y is as hereinbefore defined

R⁷ and R⁸ are each independently H, alkyl, halogen, hydroxy, amino,alkylamino, dialkylamino, alkoxy, alkylthio, cyano, nitro, ortrifluoromethyl

with the proviso that Y is not NHC(O)Me and when B is a thiophene ringthen Y is not C(O)CF₃ and when B is a 5(6)-benzimidazole ring then Y isnot NHCO₂Me or NHCO₂Et

and the pharmaceutically acceptable salts, solvates, hydrates andprodrugs thereof.

As used herein the term “alkyl”, alone or in combinations, means astraight or branched-chain alkyl group containing from one to seven,preferably a maximum of four, carbon atoms such as methyl, ethyl,propyl, isopropyl, butyl, sec-butyl, t-butyl and pentyl. Examples ofalkoxy groups are methoxy, ethoxy, propoxy, isopropoxy, butoxy andt-butoxy. The term “halogen” means fluorine, chlorine, bromine oriodine. The term “aryl” as used herein unless otherwise stated includesreference to a C₆₋₁₀ aryl group which may, if desired, carry one or moresubstituents selected from halogeno, alkyl, haloalkyl, alkoxy, hydroxy,amino, nitro and cyano. The term “aralkoxy” means an alkoxy groupsubstituted with an aryl group.

The term heteroaryl is defined herein as a mono- or bi-cyclic aromaticgroup containing one to four heteroatoms selected in any combinationfrom N, S or O atoms and a maximum of 9 carbon atoms. Examples ofheteroaryl groups include pyridyl, pyrimidyl, furyl, thienyl, pyrrolyl,pyrazolyl, indolyl, benzofuryl, benzothienyl, benzothiazolyl, oxazolyl,isoxazolyl, thiazolyl, isothiazolyl, imidazolyl, triazolyl, quinolyl andisoquinolyl groups.

The term heterocycloalkyl includes heterocycloalkyl groups containing3-6 carbon atoms and one or two oxygen, sulphur or nitrogen atoms.Particular examples of such groups include azetidinyl, pyrrolidinyl,piperidinyl, homopiperidinyl, piperazinyl, homopiperazinyl, morpholinylor thiomorpholinyl groups.

The term cycloalkyl means a cycloaliphatic group containing 3-10 carbonatoms such as, for example, cyclopropyl, cyclobutyl, cyclopentyl orcyclohexyl.

One particularly preferred group of compounds are those of formula II inwhich Y is a phosphate group.

Another particularly preferred group of compounds are those of formulaII in which Y is a group NR⁵C(O)R⁴, R⁵ is hydrogen and R⁴ is a1-aminoalkyl group which can be further substituted for example by ahydroxy group.

Where one or more functional groups in compounds of formula I or II aresufficiently basic or acidic the formation of salts is possible.Suitable salts include pharmaceutically acceptable salts for exampleacid addition salts including hydrochlorides, hydrobromides, phosphates,sulphates, hydrogen sulphates, alkylsulphonates, arylsulphonates,acetates, benzoates, citrates, maleates, fumarates, succinates, lactatesand tartrates, salts derived from inorganic bases including alkali metalsalts such as sodium or potassium salts, alkaline earth metal salts suchas magnesium or calcium salts, and salts derived from organic aminessuch as morpholine, piperidine or dimethylamine salts.

Those skilled in the art will recognise that compounds of formulae I andII may exist as stereoisomers and/or geometrical isomers and accordinglythe present invention includes all such isomers and mixtures thereof.The substituted benzimidazole group is capable of existing in tautomericforms and the formulae I and II are intended to represent all tautomericforms and the substituent AX- is in the 5(6) position.

Compounds of Formula I or II may be prepared by any process known to aperson skilled in the art. Compounds of Formula I or II may be preparedby a number of processes as generally described below and morespecifically in the Examples hereinafter. In the following processdescription, the symbols A, X and alk when used in the formulae depictedare to be understood to represent those groups described above inrelation to formula I or II unless otherwise indicated. In the schemesdescribed below it may be necessary to employ protecting groups whichare then removed during the final stages of the synthesis. Theappropriate use of such protecting groups and processes for theirremoval will be readily apparent to those skilled in the art.

Thus according to a further aspect of the invention compounds offormulae I and II in which R¹ and R² are hydrogen may be prepared bytreatment of a diamine of formula III with a1,3-bis(alkoxycarbonyl)-S-alkyl isothiourea, for example1,3-bis(methoxycarbonyl)-S-methyl isothiourea or1,3-bis(ethoxycarbonyl)-S-methyl isothiourea, in a solvent such as analcohol, for example methanol or ethanol, optionally mixed with water,at from about room temperature to the reflux temperature of the solventfor about 5 minutes to 6 hours. The reaction medium is preferably madeacidic by the addition of for example an organic acid such as aceticacid or p-toluenesulphonic acid.

Compounds of formula III are either known or can be prepared byconventional procedures.

Compounds of formulae I and II can also be prepared from other compoundsof formulae I and II by chemical modification. Example of such chemicalmodifications that may be applied are standard alkylation, acylation,reduction, oxidation, sulphation, aromatic halogenation, aromaticnitration, phosphorylation, hydrolysis, condensation, cleavage andcoupling reactions. These reactions may be used to add new substituents,to modify existing substituents or to modify other parts of themolecule.

Thus for example a compound of formula I or II in which R¹ is hydrogencan be converted into the corresponding compounds where R¹ isalkylaminocarbonyl by treatment with an alkyl isocyanate in a solventsuch as tetrahydrofuran at a temperature in the range 0° to 40° C.,typically room temperature.

In another general example a thioether group in a compound of formula Ior II can be converted into a sulphinyl group by treatment withperiodate in an aqueous alcohol solvent such as aqueous methanol or inaqeuous acetonitrile at about −20° to 50° C., for about 1 to 16 h.Alternatively this conversion can be effected by treatment with oneequivalent of a peracid such as 3-chloroperbenzoic acid in a chlorinatedsolvent such as dichloromethane or chloroform, at a temperature of about−30° C. to room temperature.

In a further general example a thioether group in a compound of formulaI or II can be converted into a sulphonyl group by treatment with two ormore equivalents of a peracid such as 3-chloroperbenzoic acid in achlorinated solvent such as dichloromethane or chloroform, at atemperature of about −30° C. to room temperature.

In a further general example a keto group in a compound of formula I orII can be converted into a carbinol group by treatment with a reducingagent for example a hydride such as lithium aluminium hydride in anether solvent such as diethyl ether or tetrahydrofuran at a temperatureof from about 0° to the reflux temperature of the solvent.

In a further general example a keto group in a compound of formula I orII can be converted into an imine by treatment with ammonia in analcoholic solvent such as ethanol at around room temperature for anextended period, for example three weeks.

In a further general example a keto group in a compound of formula I orII can be converted into an oxime by treatment with hydroxylamine in analcoholic solvent such as ethanol at around room temperature to aroundthe reflux temperature of the solvent.

In a further general example a compound of formula I or II containing ahydroxy group can be converted into the correspondingdihydrogenphosphate ester by treatment with for example di-tert-butyldiethylphosphoramidite in the presence of a suitable catalyst forexample tetrazole in a solvent such as an ether solvent for exampletetrahydrofuran at a temperature in the range −40 to 40° C.,conveniently at or near room temperature, followed by treatment with anoxidising agent for example 3-chloroperoxy benzoic acid or magnesiummonoperoxyphthalate at a temperature in the range −78° C. to 40° C.preferably −65 to −10° C. The resulting intermediate phosphate triesteris treated with an acid for example trifluoroacetic acid in a solventsuch as a chlorinated solvent e.g. dichloromethane at a temperature inthe range −30 to 40° C. conveniently at or near 0° C. to give thecompound of formula I or II containing a dihydrogenphosphate-ester.

In another general example an O-alkyl group may be cleaved to thecorresponding alcohol (OH) by reaction with boron tribromide in asolvent such as a chlorinated solvent e.g. dichloromethane at a lowtemperature e.g. around −78° C.

In a further general example compounds of formula I or II may bealkylated by reaction with a suitable alkylating agent such as an alkylhalide, an alkyl toluenesulphonate, an alkyl methanesulphonate or analkyl triflate. The alkylation reaction can be carried out in thepresence of a base for example an inorganic base such as a carbonatee.g. caesium or potassium carbonate, a hydride such as sodium hydride oran alkoxide such as potassium t-butoxide in a suitable solvent such asan aprotic solvent e.g. dimethylformarnide or an ether solvent such astetrahydrofuran at a temperature of around −10 to 80° C.

In a further general example compounds of formula I or II containing anamine group may be acylated by treatment with a carboxylic acid and acoupling reagent, for example dicyclohexylcarbodiimide, in a sutablesolvent for example an aprotic solvent such as dimethylformamide, anether solvent such as tetrahydrofuran, a chlorinated solvent for exampledichloromethane or a solvent mixture at a temperature in the range 0° to60°, preferably about room temperature.

In a further general example a compound of formula I or II containing anOH group can be converted into a carbamate by reaction with an alkylisocyanate or a carbamoyl chloride in an aprotic solvent such asdimethylfornamide, an ether solvent such as tetrahydrofuran, achlorinated solvent for example dichloromethane or a solvent mixture inthe presence of a base such as a tertiary amine base for exampletriethylamine at a temperature in the range −20° to the refluxtemperature of the solvent, conveniently at or around room temperature.

In a further general example a compound of formula I or II containing anamino group can be converted into a urea by reaction with an isocyanateor a carbamoyl chloride in an aprotic solvent such as dimethylformamide,an ether solvent such as tetrahydrofuran, a chlorinated solvent forexample dichloromethane or a solvent mixture in the presence of a basesuch as a tertiary amine base for example triethylamine at a temperaturein the range −20° C. to the reflux temperature of the solvent,conveniently at or around room temperature.

In a further general example a compound of formula I or II containing ahydroxy group can be converted into a carbonate by reaction with anchloroformnate in an aprotic solvent such as dimethylformamnide, anether solvent such as tetrahydrofuran, a chlorinated solvent for exampledichloromethane or a solvent mixture in the presence of a base such as atertiary amine base for example triethylamine at a temperature in therange −20° C. to the reflux temperature of the solvent, preferably at oraround 0° C.

Preparation of a compound of formula I or II as a single enantiomer or,where appropriate, diastereomer may be effected by synthesis from anenantiomerically pure starting material or intermediate or by resolutionof the final product in a conventional manner.

Acid addition salts of the compounds of formula I or II are prepared ina conventional manner by treating a solution or suspension of the freebase I or II with about one equivalent of a pharmaceutically acceptableacid. Salts of compounds of formula I or II derived from inorganic ororganic bases are prepared in a conventional manner by treating asolution or suspension of the free acid I or II with about oneequivalent of a pharmaceutically acceptable organic or inorganic base.Alternatively both acid addition salts and salts derived from bases maybe prepared by treatment of the parent compound with the appropriateion-exchange resin in a standard fashion. Conventional concentration andrecrystallisation techniques are employed in isolating the salts.

Compounds according to the invention are able to destroy vasculaturethat has been newly formed, for example tumour vasculature, whileleaving unaffected normal, mature vasculature. The ability of thecompounds to act in this way may be determined by the tests describedhereinafter.

The compounds according to the invention are thus of particular use inthe prophylaxis and treatment of cancers involving solid tumours and inthe prophylaxis and treatment of diseases where inappropriateangiogenesis occurs such as diabetic retinopathy, psoriasis, rheumatoidarthritis, atherosclerosis and macular degeneration.

The compounds of the invention may be administered as a sole therapy orin combination with other treatments. For the treatment of solid tumourscompounds of the invention may be administered in combination withradiotherapy or in combination with other anti-tumour substances forexample those selected from mitotic inhibitors, for example vinblastine,paclitaxel and docetaxel; alkylating agents, for example cisplatin,carboplatin and cyclophosphamide; antimetabolites, for example5-fluorouracil, cytosine arabinoside and hydroxyurea; intercalatingagents for example adriamycin and bleomycin; enzymes, for exampleaspariginase; topoisomerase inhibitors for example etoposide, topotecanand irinotecan; thymidylate synthase inhibitors for example raltitrexed;biological response modifiers for example interferon; antibodies forexample edrecolomab and antibodies against the EGFr, HER2 receptor orVEGF receptor: and anti-hormones for example tamoxifen. Such combinationtreatment may involve simultaneous or sequential application of theindividual components of the treatment.

For the prophylaxis and treatment of disease the compounds according tothe invention may be administered as pharmaceutical compositionsselected with regard to the intended route of administration andstandard pharmaceutical practice. Such pharmaceutical compositions maytake a form suitable for oral, buccal, nasal, topical, rectal orparenteral administration and may be prepared in a conventional mannerusing conventional excipients. For example for oral administration thepharmaceutical compositions may take the form of tablets or capsules.For nasal administration or administration by inhalation the compoundsmay be conveniently delivered as a powder or in solution. Topicaladministration may be as an ointment or cream and rectal administrationmay be as a suppository. For parenteral injection (includingintravenous, subcutaneous, intramuscular, intravascular or infusion) thecomposition may take the form of, for example, a sterile solution,suspension or emulsion.

The dose of a compound of the invention required for the prophylaxis ortreatment of a particular condition will vary depending on the compoundchosen, the route of administration, the form and severity of thecondition and whether the compound is to be administered alone or incombination with another drug. Thus the precise dose will be determinedby the administering physician but in general daily dosages may be inthe range 0.001 to 100 mg/kg preferably 0.1 to 50 mg/kg.

BIOLOGICAL ACTIVITY

The following test was used to demonstrate the activity and selectivityof compounds according to the invention.

Activity Against Tumour Vasculature Measured by Fluorescent Dye

The following experiment demonstrates the ability of the compounds todamage tumour vasculature.

Tumour functional vascular volume in CaNT tumour-bearing mice wasmeasured using the fluorescent dye Hoechst 33342 according to the methodof Smith et al (Brit J Cancer 57, 247-253, 1988). The fluorescent dyewas dissolved in saline at 6.25 mg/ml and injected intravenously at 10mg/kg 6 hours or 24 hours after intraperitoneal drug treatment. Oneminute later, animals were killed and tumours excised and frozen; 10 μmsections were cut at 3 different levels and observed under UVillumination using an Olympus microscope equipped with epifluorescence.Blood vessels were identified by their fluorescent outlines and vascularvolume was quantified using a point scoring system based on thatdescribed by Chalkley, (J Natl Cancer Inst, 4, 47-53, 1943). Allestimates were based on counting a minimum of 100 fields from sectionscut at the 3 different levels. Results are expressed as percentagereduction in vascular volume compared to control.

The activity of compounds of the invention in this assay is shown inTable 1.

TABLE 1 Reduction in tumour vascular volume measured by fluorescent dyeDose (mg/kg) Time % Reduction in Compound (i.p.) (h) vascular volumeFenbendazole 500  6 44 Mebendazole 500  6 56 Albendazole 500  6 51Oxibendazole 100  6 43 Nocodazole 100 24 23 Compound of Example: 1 500 6 81 2 500  6 80 3  50 24 22 4  50 24 53 5  50 24 99 12  50 24 51 14 50 24 39 15  50 24 61 16  50 24 56 17  50 24 62 18  50 24 47 19  50 2488 20  50 24 69 21  50 24 74

The following non-limiting Examples illustrate the invention:

EXAMPLE 1 Methyl [5(6)-(4-Hydroxvbenzoyl)-1H-benzimidazol-2yl]carbamate

A solution of 3,4-diamino-4′-hydroxybenzophenone (49.6mg, 0.21mmol) and1,3-bis(methoxycarbonyl)-S-methyl isothiourea (98 mg, 0.44 mmol) inethanol (2.5 ml) was treated with p-toluenesulphonic acid (7 mg) and themixture heated at reflux for 10 minutes. The mixture was cooled and theprecipitate collected by filtration and washed with ethanol and hexaneto give the title compound (16 mg) as a white solid m.p. >258° C.,¹H-NMR (400 MHz, d6-DMSO) δ 7.80 (s, 1H), 7.65 (d, 2H, J=8 Hz), 7.50 (s,1H), 7.49 (s, 1H), 6.89 (d, 2H, J=8 Hz), 3.77 (s, 3H) ppm. m/e 311 (M+).Anal. Calculated for C16H13N3O4: C, 61.73; H, 4.21; N 13.49. Found: C,61.68; H, 4.18; N 13.36.

EXAMPLE 2 Methyl[5(6)-(4-Phosphonooxybenzoyl)-1H-benzirnidazol-2yl]carbamate

A solution of methyl[5-(4-(di-tert-butylphosphonooxy)benzoyl)-1H-benzimidazol-2yl]carbamate(1.5 g, 3.0 mmol) in dichloromethane (55 ml) was cooled in an ice bathand treated with trifluoroacetic acid (6 ml) dropwise. The mixture wasallowed to warm to room temperature and stirred for 1 hour beforesolvents were removed under reduced pressure. The residue was trituratedwith ether to give the title compound (1.13 g) as a white solidm.p.>258° C., H-NMR (300 MHz, d6-DMSO) δ :7.84 (s, 1H), 7.75 (d, 1H, J=9Hz), 7.57 (d, 1H, J=8 Hz), 7.52 (d, J=8 Hz), 7.33 (d, 1H, J=9 Hz), 3.78(s, 3 H) ppm.

The methyl[5(6)-(4-(di-tert-butylphosphonooxy)benzoyl)-1H-benzimidazol-2yl]carbamateused as starting material was prepared as follows:

A solution of methyl[5(6)-(4-hydroxybenzoyl)-1H-benzimnidazol-2yl]carbamate (100 mg, 0.3mmol) in anhydrous tetrahydrofuran (1 ml) stirred under a nitrogenatmosphere was treated with di-tert-butyl diethylphosphoramidite (74 mg,0.29 mmol) and 1H-tetrazole (54 mg, 0.78 mmol) and the mixture stirreduntil the reaction was shown to be complete by TLC (about 1 h). Thecooled (−40° C.) mixture was treated with 3-chloroperbenzoic acid (79mg, 0.39 mmol) in dichloromethane (1 ml) and stirred 10 minutes beforebeing allowed to warm to room temperature. The mixture was washed withsaturated aqueous sodium bicarbonate followed by brine and the dried(MgSO4) organic phase concentrated under reduced pressure. The residuewas chromatographed on silica gel eluting first with 4%methanol/dichloromethane then with 5% methanol/dichloromethane. Methyl[5(6)-(4-(di-tert-butylphosphonooxy)benzoyl)-1H-benzimidazol-2yl]carbamate(35 mg) was obtained as a white solid m.p. 99-101° C.

EXAMPLE 3 Methyl[5(6)-(4-Phosphonooxyphenylthio)-1H-benzimidazol-2yl]carbamate

A solution of methyl[5(6)-(4-(di-tert-butylphosphonooxy)phenylthio)-1H-benzimidazol-2yl]carbamate(180 mg) in anhydrous dichloromethane (10 ml) at 4° C. was treated withtrifluoroacetic acid (1 ml) and stirred for 1.5 h. The mixture wasallowed to warm to room temperature and concentrated under reducedpressure. Ethyl acetate was added and the mixture concentrated again.The residue was triturated with diethyl ether, washed with diethyl etherfollowed by acetone/water 9:1 and dried to give the title compound (134mg) as a white solid m.p. 190-193° C. Anal. Calculated forC₁₅H₁₄N₃O₆PS.3H₂O: C, 40.1; H, 4.5; N 9.3. Found: C, 40.1; H, 3.9; N9.2.

The methyl[5(6)-(4-(di-tert-butylphosphonooxy)phenylthio)-1H-benzimidazol-2yl]carbamateused as starting material was prepared as follows:

A solution of methyl[5-(4-hydroxyphenylthio)-1H-benzimidazol-2yl]carbamate (200 mg) in amixture of anhydrous dimethylformamide (2 ml) and anhydroustetrahydrofuran (2 ml) was treated with di-tert-butyldiethyiphosphoramidite (350 mg) and the mixture stirred for 48 h at roomtemperature. The mixture was cooled to −65° C. and treated graduallywith magnesium monoperoxyphthalate (850 mg) so that the temperatureremained below −50° C. A saturated aqueous solution of sodiumbicarbonate was added, keeping the temperature below −40° C. during theaddition then allowing the mixture to warm to room temperature. Themixture was extracted with three portions (50 ml each) of ethyl acetateand the combined extracts washed with brine (50 ml), dried (MgSO4) andconcentrated under reduced pressure. The residue was purified by radialchromatography on silica gel eluting with dichloromethane/methanol 9:1to give methyl[5(6)-(4-(di-tert-butylphosphonooxy)phenylthio)-1H-benzimidazol-2yl]carbamate(180 mg)as a white foam.

EXAMPLE 4 Methyl [5(6)-(4-Aminophenylthio)-1H-benzimidazol-2yl]carbamate

Methyl [5(6)-(4-(acetylamino)phenylthio)-1H-benzimidazol-2yl]carbamate(602 mg, 1.78 mmol) was dissolved in mixture of methanol (24 ml) andhydrochloric acid (10%, 6 ml) and heated under reflux for 16 h. Thesolution was neutralised with ammonia solution and the methanol removedunder reduced pressure. The white precipitate was collected byfiltration, washed with water and dried in vacuo to give 392 mg of apale yellow solid m.p. 282-284° C. m/e 298 (M⁺).

EXAMPLE 5 Methyl[5(6)-(4-Alanylaminophenyjthio)-1H-benzimidazol-2yl]carbamate

A suspension of methyl[5(6)-(4-(Nα-tert-butoxycarbonylalanylamino)phenylthio)-1H-benzimidazol-2yl]carbamate(250 mg) in dichloromethane (20 ml) was treated with trifluoroaceticacid (4 ml). The mixture was allowed to warm to room temperature andconcentrated under reduced pressure. Ethyl acetate was added and themixture concentrated again. The residue was triturated with diethylether to afford the trifluoroacetic acid salt of the title compound (105mg) as a white solid m.p. 178-182° C. m/e 485 (M+). Anal. Calculated forC₁₈H₁₉N₅O₃S.2C₂HF₃O₂ C; 43.1, H; 3.5, N; 11.4 Found C; 42.8, H; 3.8, N;11.3.

The methyl[5(6)(4-(Nα-tert-butoxycarbonylalanylamino)phenylthio)-1H-benzimidazol-2yl]carbamateused as starting material was prepared as follows:

A suspension of methyl[5(6)-(4-aminophenylthio)-1H-benzimidazol-2yl]carbamate (150 mg) inanhydrous tetrahydrofuran (4 ml) was treated withN-tert-butoxycarbonylalanine (100 mg), cooled to −35° C. and treatedwith 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (100 mg). The mixturewas allowed to warm to room temperature and stir for 16 h. Water (40 ml)and ethyl acetate (20 ml) were added and the mixture extracted with fourportions of ethyl acetate (50 ml each). The combined extracts werewashed with brine (50 ml), dried (MgSO4) and concentrated under reducedpressure. The residue was purified on silica gel eluting with ethylacetate to give methyl[5(6)-(4-(Nα-teri-butoxycarbonylalanylamino)phenylthio)-1H-benzimidazol-2yl]carbamate(258 mg) as a white solid m.p. 222-224° C. m/e 485 (M⁺).

EXAMPLE 6 Methyl[5(6)-(4-(Methoxycarbonylamino)phenylthio)-1H-benzimidazol-2yl]carbamate

Methyl [5(6)-(4-aminophenylthio)-1H-benzimidazol-2yl]carbamate(150 mg,048 mmol) was dissolved in dimethylformamide (2 ml) and tetrahydrofuran(2 ml) and methyl chloroformate (40 mg, 0.42 mmol) added together withtriethylamine (43 mg, 0.42 mmol). The solution was stirred at 20° C. for72 h and then evaporated to dryness under reduced pressure. The residuewas purified by chromatography on silica gel eluting with 5%methanol/dichloromethane to give the title compound as a white solid;mp>350° C. (dec.).

EXAMPLE 7 Methyl[5(6)4-(Phenylaminocarbonylamino)phenylthio)-1H-benziridazol-2yl]carbamate

Methyl [5(6(4-aminophenylthio)-1H-benzimidazol-2yl]carbamate (150 mg,0.48 mmol) was dissolved in dimethylformamide (2 ml) and tetrahydrofiran(2 ml) and phenylisocyanate (56.8 mg, 0.48 mmol) added together withtriethylarnine (50 mg, ca.0.5 mmol). The solution was stirred at 20° C.for 12 h and the solvents removed in vacuo. The residue was purified onsilica (ethyl acetateihexane, 2:1) to give the title compound as a whitesolid; mp 335-340° C. (dec.).m/e 433 (M⁺).

EXAMPLE 8 Methyl[5(6)4-(Methoxycarbonyloxn)benzoyl)-1H-benzimidazol-2yl]carbamate

Methyl [5(6)-(4-hydroxybenzoyl)-1H-benzimidazol-2yl]carbamate (110 mg,0.35 mmol) was dissolved in dimethylformamide (3.5 ml) andtriethylarnine (0.5 ml). The solution was cooled to 0° C. and methylchloroformate (50 mg, 0.52 mmol) added with stirring. The solution wasstirred for 0.5 h at 0° C. and the 1 h at 20° C. and evaporated todryness. The residue was dissolved in ethyl acetate (50 ml) and washedwith sodium bicarbonate (sat., aq., 50 ml) and brine (50 ml), dried andevaporated. The residue was purified by radial chromatography on silicagel eluting with ethyl acetate/hexane, 1:1 followed by ethyl acetate togive the title compound as a white solid; mp 224-226° C. (dec).m/e 369(M⁺).

Prepared in an analogous fashion to Example 1 were:

EXAMPLE 9

Methyl [5(6)-(2-Methoxycarbonylphenylthio)-1H-benzimidazol-2yl]carbamatefrom methyl 2-(3,4-diarminophenylthio)benzoate (1.5 g) and1,3-bis(methoxycarbonyl)-S-methyl isothiourea (2.25 g) there wasobtained the title compound (1.51 g) as a white solid m.p. 228-230 m/e357 (M+). Anal. Calculated for C₁₇H₁₅N₃O₄S: C, 57.13; H, 4.23; N 11.75.Found: C, 57.39; H, 4.13; N 11.81.

EXAMPLE 10

Methyl [5(6)-(3-Methoxvcarbonylphenylthio)-1H-benzimidazol-2yl]carbamatefrom methyl 3-(3,4-diaminophenylthio)benzoate (246 mg) and1,3-bis(methoxycarbonyl)-S-methyl isothiourea (371 mg) there wasobtained the title compound (183 mg) as an off-white solid m.p. 226-228m/e 357 (M+). Anal. Calculated for C₁₇H₁₅N₃O₄S: C, 57.13; H, 4.23; N11.75. Found: C, 56.58; H, 4.31; N 11.86.

EXAMPLE 11

Methyl [5(6)-(4-methoxycarbonylphenalthio)-1H-benzimidazol-2yl]carbamatefrom methyl 4-(3,4-diaminophenylthio)benzoate (830 mg) and1,3-bis(methoxycarbonyl)-S-methyl isothiourea (1.25 g) there wasobtained the title compound (694 mg) as an off-white solid m.p. 280-282m/e 357 (M+). Anal. Calculated for C₁₇H₁₅N₃O₄S: C, 57.13; H, 4.23; N11.75. Found: C, 57.21; H, 4.31; N 11.73.

EXAMPLE 12

Methyl [5(6)-(4-hydroxyphenylthio)-1H-benzimidazol-2yl]carbamate from4-(4-hydroxyphenylthio)-1,2-phenylenediamine (4 g) and1,3-bis(methoxycarbonyl)-S-methyl isothiourea (6 g) there was obtainedthe title compound (2.4 g) as a white solid m.p. 202-204° C. m/e 315(M⁺).¹H-NMR (400 MHz, d6-DMSO) δ 3.74 (s, 3H,), 6.76 and 7.2 (AA′BB′,4H, J=8.6 Hz), 7.03 (dd, 1H, J=1.7, 6.6 Hz,), 7.29 (d, 1H, J=1.4 Hz,),7.34 (d, 1H, J=8.3 Hz,), 9.68 (b, 1H,), 11.67 (b, 2H) ppm.

EXAMPLE 13

Methyl [5(6)-(4-(acetylamino)phenoxy)-1H-benzimidazol-2yl]carbamate from4-4-(acetylamino)phenoxy)-1,2-phenylenediamine (1.02 g) and1,3-bis(methoxycarbonyl)-S-methyl isothiourea (2.67 g) there wasobtained the title compound (0.93 g) as a white solid m.p. 304-306° C.m/e 340 (M⁺). Anal. Calculated for C₁₇H₁₆N₄O₄: C, 60.00; H, 4.74; N16.46. Found: C, 60.03; H, 4.72; N 16.42.

Prepared in an analogous fashion to Example 3 was:

EXAMPLE 14

Methyl [5(6)-(4-phosphonooxyphenoxy-1H-benziniidazol-2yl]carbamate frommethyl[5(6-(4-(di-tert-butylphosphonooxy)phenoxy)-1H-benzimidazol-2yl]carbamate(200 mg) there was obtained the title compound (140 mg) as a white solidm.p. 272-275° C.

EXAMPLE 15

Methyl[5(6)-(4-hydroxy-α-hydroxyiminobenzl)-1H-benzimidazol-2yl]carbamate[5(6)-(4-hydroxybenzoyl)-1H-benzimidazol-2yl]carbamate (100 mg, 0.32mmol) was added to a solution of hydroxylamine (0.8 mmnol) in MeOH (20mL, prepared by treatment of hydroxylamine hydrochloride (0.64 g, 1.6mmol) and NaOH (0.14 g, 1.6 mmol) followed by filtration). The solutionwas heated at 70° C. for 36 h, cooled and water (30 ml) added. Thesolution was filtered and washed with ether and the solid trituratedwith methanol to yield the title compound (40 mg) as a white solid; mp288-290° C.

The following known compounds were prepared by literature methods:

Example 16: Methyl[5(6)-(4-(acetylamino)phenoxy)-1H-benzimidazol-2yl]carbamate

Example 17: Methyl [5(6)-(4-arninophenoxy)-1H-benzimidazol-2yl]carbamate

Example 18: Methyl [5(6)-(3-aminophenoxy)-1H-benzimidazol-2yl]carbamate

Example 19: [5(6)-(4-hydroxybenzoyl)-1H-benzimidazol-2yl]carbamate

Example 20: [5(6)-(2-hydroxybenzoyl)-1H-benzimidazol-2yl]carbamate

Example 21: [5(6)-(3-hydroxybenzoyl)-1H-benzimidazol-2yl]carbamate.

What is claimed is:
 1. A composition for treatment of diseases involvingangiogenesis comprising at least one 5(6)-substitutedbenzimidazole-2-carbamate of formula I in an amount sufficient to damagenew vasculature:

wherein: alk is an alkyl group; X is oxygen, sulfur, sulphinyl,sulphonyl, carbonyl (CO), thiocarbonyl (CS), sulphonyloxy, NH,iminomethylene (C═NH), N-hydroxyiminomethylene, N-alkoxyiminomethylene,dialkoxymethylene, 1,3-dioxolan-2yl, 1,1-ethenyl, a group CHR³ or abond; R¹ is hydrogen, alkylaminocarbonyl oralkoxycarbonyl; R² ishydrogen, alkoxycarbonyl, cyanomethyl, cyanoethyl, alkoxymethyl oracetoxymethyl; R³ is hydrogen, hydroxy, alkoxy or amino; A is asubstituted aromatic ring or substituted alkyl group in which at leastone substituent is Y selected from phosphate, alkylphosphate, NHCOR⁴ andNR⁵COR⁴ where R⁴ and R⁵, which may be the same or different, are each analkyl group substituted with one or more substituents selected fromhydroxy, amino, alkylamino, guanidino, carboxyl, SO₃H, sulfate andphosphate; and prodrugs and pharmaceutically acceptable salts, solvatesand hydrates thereof.
 2. A composition according to claim 1 wherein Y isNHC(O)R⁴ wherein R⁴ is a 1-aminoalkyl group.
 3. A composition accordingto claim 1 wherein Y is a phosphate group.
 4. A composition fortreatment of diseases involving angiogenesis comprising at least one5(6)-substituted benzimidazole-2-carbamate of formula II in an amountsufficient to damage new vasculature:

wherein: alk is an alkyl group; B is an aromatic ring; X is oxygen,sulfur, sulphinyl, sulphonyl, carbonyl (CO), thiocarbonyl (CS),sulphonyloxy, NH, iminomethylene (C═NH), N-hydroxyiminomethylene,N-alkoxyiminomethylene, dialkoxymethylene, 1,3-dioxolan-2yl,1,1-ethenyl, a group CHR³ or a bond; R¹ is hydrogen, alkylaminocarbonylor alkoxycarbonyl; R² is hydrogen, alkoxycarbonyl, cyanomethyl,cyanoethyl, alkoxymethyl or acetoxymethyl; R³ is hydrogen, hydroxy,alkoxy or amino; Y is selected from phosphate, alkylphosphate, NHCOR⁴and NR⁵COR⁴ where R⁴ and R⁵, which may be the same or different, areeach an alkyl group substituted with at least one substituent selectedfrom the group consisting of hydroxy, amino, alkylamino, guanidino,carboxyl, SO₃H, sulfate and phosphate; R⁷ and R⁸ are each independentlyH, alkyl, halogen, hydroxy, amino, alkylamino, dialkylamino, alkoxy,alkylthio, cyano, nitro, or trifluoromethyl; and prodrugs andpharmaceutically acceptable salts, solvates and hydrates thereof.
 5. Acomposition according to claim 4 wherein Y is NHC(O)R⁴ wherein R⁴ is a1-aminoalkyl group.
 6. A composition according to claim 4 wherein Y is aphosphate group.